Bioavailability / bioefficacy enhancing activity of cuminum cyminum and extracts and fractions thereof

ABSTRACT

The present invention relates to a bioenhancing/bioavailability-facilitating composition comprising:  
     an effective amount of an extract and/or at least one bioactive fraction from  Cuminum cyminum;    
     one or more additive selected from drugs, nutrients, vitamins, nutraceuticals, herbal drugs/products, micro nutrients, antioxidants along with pharmaceutically acceptable additives/excipient, and  
     optionally, an effective amount of piperine or extract/fraction of piper nigrum or piper longum; and  
     a process for the preparation of such extracts and active fractions from plant  Cuminum cyminum.

FIELD OF THE INVENTION

[0001] The present invention relates to a composition containing extractand/or bioactive fractions from the plant Cuminum cyminum as abioavailability enhancer. The present invention also relates to acomposition containing extract and/or bioactive fractions from the plantCuminum cyminum with piperine as a bioavailability enhancer. The presentinvention in addition relates to the use of bioavailability and/orbioefficacy enhancers—also termed as bioenhancers or BE and methods oftheir preparation which include their isolation from a natural sourceand obtaining the final products in their chemically characterized orfingerprint-profiled form.

DESCRIPTION OF RELATED ART

[0002] Several approaches have been adopted in the past to maximize oralbioavailability, such as (a) particle size reduction (micronization,nanonization, etc.,) (b) polymorphic or crystal size and form selection,(c) solubilization of lesser soluble drugs by way of chemicalmodifications, complexation and use of co-solvents/surfactants, (d)targeted delivery of drug at the site of action, (e) controlled drugdelivery by film coating or use of polymeric matrices for sustainedrelease of drugs, (f) prodrug approach, and (g) micro-encapsulationusing liposomes.

[0003] However, based on clues from Ayurvedic literature, a new approachof increasing the bioavailability of drugs including poorly-bioavailabledrugs had been conceptualized at applicants institute Regional ResearchLaboratory (RRL), Jammu. One of the groups of herbals which has beendocumented very frequently as essential part of about 70% of Ayurvedicprescriptions, was noted to be ‘Trikatu’, that comprises three acridsviz. long pepper, black pepper and dry ginger in equal proportions. Asingle major alkaloidal constituent from peppers (piperine) was found tobe responsible for bioavailability enhancing effect. The role of gingeris to regulate intestinal function to facilitate absorption. Influenceof piperine was extensively studied on anti-TB drugs. It was determinedthat in combination with pipeline, the dose of rifampicin can be reducedby about 50% while retaining the therapeutic efficacy of this anti-TBdrug at par with the standard dose (450 mg). Based on these findingsseveral other reputed plants were evaluated forbioavailability/bioefficacy enhancing activity. Polar and non-polarextracts of parts of a few plants viz., Zingiber officinale, and Carumcarvi increased significantly (25-300%), [Applicants co-pending patentapplications] the bioavailability of a number of classes of drugs, forexample, but not limited to, antibiotics, antifungals, anti-virals,anticancer, cardiovascular, CNS, anti-inflammatory/anti-arthritic,anti-TB/antileprosy, antihistaminic/respiratory disorders,corticosteroids, immunosuppressants, anti-ulcer. Such extracts either inpresence or absence of piperine have been found to be highly selectivein their bioavailability/bioefficacy enhancing action.

[0004]Cuminum cyminum (Linn.) (Umbelliferae) is a small, slender annualherb, which is grown extensively in South-Eastern Europe, North Africabordering the Mediterranean-sea, in India and China. It is cultivated inalmost all the states in India. The chief areas are reported to be U.P., Punjab, Rajasthan, Gujarat and Maharashtra. The plant prefers a mildclimate and grows from sea level up to an elevation of 10,000 feet.

[0005] Its seeds have been used as an important condiment. In Ayurvedait is documented as katu, ushna and pacifies deranged vata. It is aneffective gastric stimulant, beneficial in abdominal lump, flatulence,diarrhoea, sprue and a strong anthelmintic. It has therapeutically beenused as an anti-diarrhoeal, galactagogue, diuretic and also beneficialin hoarseness of voice. It is also used as astringent, carminative.Paste of seeds when applied externally allays pain and irritation due toworms in the abdomen. Oil is useful in eczema.

[0006] Based on our past experience with the development of piperine asbioavailability enhancer from plants which are otherwise part of humandiet and also documented to possess medicinal properties, we took upthis plant to evaluate its bioavailability enhancing effect, if any,based on its attributes bearing some similarities to the plant sourcesof piperine.

[0007] Chemistry of Cuminum cyminum

[0008] The seeds of Cuminum cyminum were analyzed and it was reportedthat, analytical contents of seeds are (in percentage): moisture 11.9;protein 18.7; ether extractive 15.0; carbohydrates 36.6; fibre 12.0;mineral matter 5.8; calcium 1.08; phosphorus 0.49%; iron 31.0 mg/100 g;carotene calculated as vitamin A 870 1.U./100 gm; and vitamin C 3.0mg/100 g (Hlth Bull., No. 23, 1941,36).

[0009] The seeds on distillation yield a volatile oil (2.0-4.0%) havingan unpleasant characteristic odour, spicy and somewhat bitter taste. Theoil is colourless or yellow when fresh, turning dark on keeping. Theanalytical constants of the Indian oil are d^(15°,) 0.8945; n_(D) ^(25°)1.490; [α]_(D) ^(25°), +3.60; aldehydes 16%; the oil is soluble in 11volume of 80% alcohol at 20° C. The ranges of constants reported byParry are: sp. gr. 0.900-0.930;

[0010] n=1.494-1.507; [α]+3.0 to +8.0°; aldehydes 25-35% (Rao et al, J.Indian Inst. Sci., 1925, 8A, 182; Parry, E. J. “The Chemistry ofEssential Oils and Artificial Perfumes” [1921] (Scott, Greenwood & SonLtd., London) Vol. 1, p.311).

[0011] The chief constituent of the volatile oil is cuminaldehyde(C₁₀H₁₂O, p-isopropylbenzaldehyde, b.p. 2350), which forms nearly 20-40%of the oil. Besides the aldehyde, oil contains p-cymene, pinene,dipentene, cumene, cuminic alcohol, β-phellandrene and α-terpineol. Theresidue left after the volatile oil extraction contains 17.2% proteinand 30.0% fat. It can be used as cattle feed (Finnemore, H. “TheEssential Oils” [1926] (Ernest Benn. Ltd., London) p. 641). Besidesvolatile oil seeds contains 10% fixed oil, which is greenish brown incolor with a strong aromatic flavor.

[0012] Other chemical constituents reported are apigenin-7-glucoside,apigenin-7-diglucoside, apigenin-7,4′-diglucoside,apigenin-7-digalacturonide, apigenin-7-galacturonylglucoside,apigenin-7-digalacturonide-4′-glucoside, apigenin-6,8-di-C-glucoside,luteolin-glucoside, luteolin-7-diglucoside, luteolin-7,3′-diglucoside,luteolin-7,4′-diglucoside and luteolin-7-galacturonide-4′-glucoside andchrysoeriol glycoside [El-Negoumy, S. I. et al. Grasas Aceites (Seville)1989, 40 (2) 87-9].

[0013] There is a great interest and medical need for the improvement ofbioavailability of a large number of drugs, which are (a) poorlybioavailable, (b) administered for long periods, (c) toxic and (d)expensive. Maximizing oral bioavailability is therapeutically importantbecause the extent of bioavailability directly influences plasmaconcentrations and consequently therapeutic efficacy and dose relatedtoxic effects resulting after oral drug administration. Poorlybioavailable drugs remain sub-therapeutic because a major portion of adose never reaches the plasma or exerts its pharmacological effectunless and until very large doses are given which may lead to seriousside effects. Any significant improvement in bioavailability will resultin lowering the dose or the dose frequency of that particular drug.Besides, inter-subject variability is inversely correlated with theextent of bioavailability. Therefore, low oral bioavailability leads tohigh variability and poor control of plasma concentration andpharmacodynamic effects. Inter-subject variability is particularly ofconcern for a drug with a narrow safety margin.

[0014] Incomplete oral bioavailability has various causes. These includepoor dissolution or low aqueous solubility, poor intestinal membranepermeation, degradation of the drug in gastric or intestinal fluids andpre-systemic intestinal or hepatic metabolism. The normal practice tooffset some of these problems has been to increase the dosage as statedearlier, which has the concerns of toxicity patients' non-compliance.

[0015] Many therapeutic treatments are also accompanied by loss ofessential nutraceuticals in the course of therapy. The present inventionimproves nutritional status by increasing bioavailability/bioefficacy ofvarious nutraceuticals also, which include metals and vitamins. Thebioenhancers of the invention also have the potential to enhance thebioefficacy of a drug without influencing its plasma concentrations forvarious reasons, some of which, but not limited to, are described laterin this invention under Section on ‘Bioavailability/Bioenhancingactivity’.

[0016] Bioavailability/Bioefficacy Enhancing Activity

[0017] In the present invention, the term Bioavailability orbioenhancing (BE) activity is defined as “a product at a lower dosagelevel which in combination with a drug or nutrient, provides moreavailability of the drug there by reducing the consumption of the drugor nutrient resulting in enhanced efficacy of the said drug.

[0018] In the present invention, the aqueous, aqueous—alcoholic,ketonic, ethereal, halogenated solvent extracts of the plant parts wereevaluated with different therapeutic categories of drugs andnutraceutical (vital amino acids, metals, antioxidants, vitamins), andherbal drugs either alone or in combination. The bioavailabilityenhancing (BE) activity of the extracts was found to be consistent from2.0 mg to 100 mg irrespective of the amount of the drug(s) present inthe formulation. Sub-fractions of the extracts were also evaluated, withthe same categories of drugs. The BE activity of the fractions increasedcorresponding to their proportions in the parent extract. The doses ofthe fractions responsible for the BE activity ranged from 0.5 to 25.0mg. Both the fractions were found to be equally active within theabove-mentioned range. Both the parent extracts as well as the fractionswere found to be active individually as well as in combination with eachother with different categories of drugs. The bioenhancer activity ofthe fraction (s) was found to be consistent from 0.5 mg to 25.0 mgirrespective of the amount of the drug(s) present in the formulation.The BE activity of the fractions was more enhanced as compared to thatof the parent extracts.

[0019] The extracts or its fractions were found to be upto 50% moreactive when used individually in combination with piperine (1-piperoylpiperidine). Besides, both the parent extracts as wells as theirfractions in different combinations showed almost similar enhancedactivity upto 60% in presence of piperine. The amount of piperine inthese formulations ranged from 3-20 mg.

[0020] The extracts or its fractions either in presence or absence ofpiperine have been found to be highly selective in their bioavailabilityenhancing activity. This is apparent from the degree of bioavailibilityenhancement caused by these extracts/fractions. It varies from Nil tonearly significant (25%) to highly significant (435%). The reasons forthis selective pattern may be attributable to one or more than one ofthe following reasons:

[0021] (a) Promoting the absorption of drugs from GIT,

[0022] (b) Inhibiting or reducing the rate of biotransformation of drugsin the liver or intestines,

[0023] (c) Modifying the immune system in a way that the overallrequirement of the drug is reduced substantially,

[0024] (d) Increasing the penetration or the entry into the pathogenseven where they become persistors within the macrophages such as forMycobacterium tuberculosis and such others. This eventually ensures theenhanced killing of these organisms well secured within the placesotherwise inaccessible to the active drug,

[0025] (e) Inhibiting the capability of pathogens or abnormal tissue toreject the drug e.g., efflux mechanisms frequently encountered withanti-malarial, anti-cancer and anti-microbial drugs,

[0026] (f) Modifying the signalling process between host and pathogenensuring increased accessibility of the drugs to the pathogens,

[0027] (g) Enhancing the binding of the drug with the target sites suchas receptors, proteins, DNA, RNA, and the like in the pathogen, thuspotentiating and prolonging its effect leading to enhanced antibioticactivity against pathogens,

[0028] (h) Besides above plausible modes of action, the bioenhanceragents may also be useful for promoting the transport of nutrients andthe drugs across the blood brain barrier, which could be of immense helpin the control of diseases like cerebral infections, epilepsy and otherCNS problems.

[0029] Primarily, but not exclusively, the invention enhances thecarrier mediated entry of drugs and also the passive diffusion and theactive transport pathways in the tissue which are responsible fortransporting physiological substances such as nutraceuticals to theirtarget sites. As applicable to any mechanism of action the products ofthis invention contribute in a synergistic and/or additive manner sothat most drugs and nutraceuticals in presence of the products describedin the present art are more bioavailable or bioefficaceous as a resultof one or more of these mechanisms. The bioavailability and/orbioefficacy of drugs and nutraceuticals is also relevant to animalhealth besides being important for humans. The invention therefore isalso intended to be used in veterinary preparations.

OBJECTS OF THE INVENTION

[0030] The main object of the invention is to provide an active ofextract and bioactive fraction obtained from Cuminum cyminum.

[0031] Another object of the invention is to evaluatebioenhancing/bioavailability of Cuminum cyminum extract or bioactivefraction in combination with drugs, nutrients, nutraceuticals,micronutrients and herbal drugs/products.

[0032] Still another object of the invention is to provide a bioenhancercomposition comprising active principles of Cuminum cyminum incombination with drugs, nutrients, nutraceuticals, micronutrients andherbal drugs/products.

[0033] Still another embodiment of the present invention is to provide abioenhancer composition comprising extract and/or bioactive fractionsobtained from Cuminum cyminum, piperine and one or more selected fromthe group consisting of drugs, nutrients, nutraceuticals, micronutrientsand herbal drugs/products.

[0034] Yet, another object of the invention is to provide a process forisolating bioactive factions from Cuminum cyminum useful as abioenhancer.

[0035] Yet, another object of the invention is to provide a process forisolating bioactive faction from Cuminum cyminum using aqueous and/oralcoholic solvent

SUMMARY OF THE INVENTION

[0036] Accordingly, the present invention is directed to preparation ofactive extracts/fraction from the plant Cuminum cyminum which includetheir chemical characterization, fingerprint profiling and methods ofusing such products to enhance bioavailability and/or bioefficacy ofdrugs, natural products and essential nutraceuticals. The presentinvention is directed to preparation of composite bioenhancerscomprising polar and non-polar extracts of parts of Cuminum cyminumand/or piperine (Ex: Piper nigrum and Piper longum) which increasedsignificantly (25 to 435%), the bioavailability of a number of classesof drugs, for example, but not limited to, antibiotics, antifungals,anti-virals, anticancer, cardiovascular, CNS,anti-inflammatory/anti-arthritic, anti-TB/anti leprosy,anti-histaminic/respiratory disorders, corticosteroids,immunosuppressants, anti-ulcer. Such extracts/fractions of C. cyminumeither in presence or absence of piperine (Ex: Piper nigrum and Piperlongum) have been found to be highly selective in theirbioavailability/bioefficacy enhancing action.

BRIEF DESCRIPTION OF THE ACCOMPANYING DRAWINGS

[0037]FIG. 1 represents HPLC chromatogram of aqueous extract of Cuminumcyminum

[0038]FIG. 2 represents HPLC chromatogram of 50% aqueous alcoholicextract of Cuminum cyminum

[0039]FIG. 3 represents HPLC chromatogram of fraction 1 of Cuminumcyminum

[0040]FIG. 4 represents HPLC chromatogram of fraction 2 of Cuminumcyminum

[0041]FIG. 5 represents HPLC chromatogram of fraction 3 of Cuminumcyminum

[0042]FIG. 6 represents HPLC chromatogram of fraction 4 of Cuminumcyminum

[0043]FIG. 7 represents HPLC chromatogram of fraction 5 of Cuminumcyminum

DETAILED DESCRIPTION OF THE INVENTION

[0044] Accordingly, the present invention provides abioenhancing/bioavailability-facilitating composition comprising:

[0045] i. an effective amount of an extract and/or at least onebioactive fractions from Cuminum cyminum;

[0046] ii. one or more additive selected from drugs, nutrients,vitamins, nutraceuticals, herbal drugs/products, micro nutrients,antioxidants along with pharmaceutically acceptable additives/excipient,and

[0047] iii. optionally, an effective amount of piperine orextract/fraction of piper nigrum or piper longum.

[0048] One embodiment of the invention the amount of Cuminum cyminumextract used is in the range of about 1.0 to 250 mg, preferably in therange of 2.0 to 100 mg.

[0049] Another embodiment, the amount of Cuminum cyminum fraction usedis in the range of about 0.5 to 75.0 mg irrespective of the amount ofadditive selected from drugs, nutrients, vitamins, nutraceuticals,herbal drugs/products, micro nutrients and antioxidants, preferably inthe range of about 1.0 to 30 mg. The fraction Cuminum cyminum used isselected from fractions identified as 1 to 5 The amount of piperine usedis in the range of about 3 to 50 mg, more particularly between 3 to 20mg, wherein the piperine is isolated from piper nigrum, piper longum orits oleoresin.

[0050] Another embodiment of the invention provides a composition inwhich the drugs are selected from the group consisting of antibiotics,antifungal, antiviral, anticancer, cardiovascular, CNS drugs,anti-inflammatory/anti-arthritic, anti-TB/antileprosy drugs,anti-histamines/drugs for respiratory disorders, corticosteriods,immuno-suppressants, anti-ulcer drugs and herbal drugs.

[0051] Still another embodiment, the antibiotic used is selected fromthe group consisting of quinolones, macrolides, cephalosproins,penicillin and aminoglycosides; the quinolone is selected from the groupconsisting of Ciprofloxacin, Pefloxacin, Ofloxacin and Norfloxacin; themacrolide is selected from the group consisting of Erythromycin,Roxythromycin and Azithromycin; the cephalosproins is selected from thegroup consisting of Cefalexin, cefatrioxone, cefixime, Cefpirome,Cefdinir and Cefadroxil; the penicillin is selected from the groupconsisting of Amoxycillin and Cloxacillin; and the aminoglycocide isselected from the group consisting of Amikacin and Kanamycin.

[0052] Still another embodiment, the anti-fungal drug is selected fromthe group consisting of Fluconazole, Amphotericin B, Griseofulvin andKetoconazole and the antiviral drug is selected from the groupconsisting of Acyclovir and Zidovudine.

[0053] Yet another embodiment, the anticancer drug is selected from thegroup consisting of Methotrexate, 5-Fluorouracil, Doxorubicin, Taxol andCisplatin.

[0054] Another embodiment, the cardiovascular drug is selected from thegroup consisting of Amlodipin, Lisinopril, propranolol and Atenolol andCNS drugs is selected from the group consisting of Alprazolam andHaloperidol Still another embodiment anti-inflammatory/anti-arthriticdrug is selected from the group consisting of Diclofenac, Piroxicam,Nimesulide and Rofecoxib and anti-TB/anti-leprosy drug is selected fromthe group consisting of Rifampicin, Ethionamide, Isoniazid, Cycloserine,Pyrazinamide, Ethambutol and Dapsone

[0055] The antihistamine/drugs for respiratory disorders compound isselected from the group consisting of Salbutamol, Theophylline,Bromhexine and Loratidine; the corticosteriod is selected from the groupconsisting of Prednisolone, dexamethasone and Betamethasone;immuno-supressant is selected from the group consisting of CyclosporinA, Tacrolimus and Mycophenolatemofetil and the anti-ulcer compound isselected from the group consisting of Rantidine, Cimetidine andOmerprazole.

[0056] Yet another embodiment the herbal product/drug is selected fromEchinacea, Tinospora cordifolia, Picrorrhiza kurroa, Aegles marmelos,Andrographis paniculata, Emblica ribes, Asparagus racemosus, Terminaliachebula Withania somnifera, Centella asiatica and/or their mixturethereof.

[0057] Another embodiment of the invention relates to a composition,wherein the nutrient is selected from group consisting of sugar,carbohydrates, fats and proteins, the vitamin is selected from the groupconsisting of Vitamin A, Vitamin E, Vitamin B1, Vitamin B6, Vitamin B12,Vitamin C and Folic acid; and the antioxidant is selected from the groupconsisting of β-Carotene, Silymarin, Selenium, Lycopene andEllagiogallotannins Still another embodiment, natural herbal product isselected from the group consisting of Curcumin, Boswellic acids andRutin and essential micro nutrients is selected from the groupconsisting of Methionine, Lysine, Leucine, Valine, Isoleucine, Zinc,Calcium, Glucose, Potassium, Copper and Iron

[0058] Yet another embodiment, the plant extract of Cuminum cyminum orits bioactive fraction is extracted from any plant parts of Cuminumcyminum

[0059] In another embodiment, the above said composition is administeredthrough oral, parenteral, nasal, inhalation including nebulisers,rectal, vaginal, transdermal and any others suitable routes.

[0060] In another embodiment, the bioenhancing effect of theextracts/fractions of Cuminum cyminum alone or in combination withpiperine is selective in enhancing the bioavailability/bioefficacy of adrug, nutraceutical, and herbal drug/formulation.

[0061] One more embodiment of the invention provides the compositioncontaining Cuminum cyminum extract or the fractions there of whichprovides bioavailability/bioenhancing activity in the range of 25-335%

[0062] Still another embodiment, the composition comprising componentsof Cuminum cyminum and piperine exhibit nearly significant i.e. about25% to highly significant i.e. about 435% bioenhancing/bioavailabilityactivity.

[0063] Yet another embodiment, the said composition-containing piperineand Cuminum cyminum provides further bioavailability/bio-enhancingactivity in the range of 10 to 85% more than bioenhancing activity ofCuminum cyminum alone.

[0064] Another embodiment of the invention relates to the dosage levelof the composition comprising Cuminum cyminum extract is in the range of10 to 30-mg/kg/body weight and composition comprising bioactive fractionis in the range of 2 to 20-mg/kg/body weight.

[0065] Another embodiment of the invention relates to the dosage levelof the composition comprising Cuminum cyminum extract or bioactivefraction along with piperine, wherein the dosage of piperine is in therange of 0 to 12-mg/kg/body weight.

[0066] One more embodiment of the present invention provides a processfor the preparation of an aqueous extract, aqueous alcoholic extract andbioactive fraction from the plant Cuminum cyminum, said processcomprises steps of:

[0067] a) extracting crushed plant material with water or 50% aqueousalcoholic solvent at a temperature range of 95-100° C. to obtain aq.extract or aq. alcoholic extract respectively,

[0068] b) extracting a portion of aq. extrtact of step (a) withn-butanol (n-BuOH), separating the n-butanol layer and a aqueous layer,

[0069] c) evaporating, freeze drying n-BuOH layer of step (b) to obtainfraction 1,

[0070] d) evaporating, freeze drying the aq. layer of step (b) to obtainfraction 2,

[0071] e) refluxing another portion of the aq.layer of step (a) withalcohol, pooling the alcohol extract and separating the residue leftover,

[0072] f) evaporating the pooled alcohol extract of step (e) to obtain aresidue as fraction 3,

[0073] g) extracting residue of step (e) with 50% aq.alcoholic solventto obtain 50% aq.alcoholic soluble portion and residue as fraction 5,and

[0074] h) evaporating the aqueous alocoholic soluble portion of step (g)to obtain a residue as fraction 4.

[0075] One more embodiment of the invention provides HPLC chromatogramfor the extract and bioactive fractions obtained from plantCuminum.cyminum.

[0076] Still another embodiment, the HPLC chromatrogaph is obtained byusing 2% acetic acid in water:acetonitrile in the ratio 83:17, RP18column; flow rate 1 ml/minute using UV detector.

[0077] Another embodiment, the aqueous extract obtained from the plantCuminun cyminum is having HPLC chromatograph major peaks with theretention time 2.16, 2.44, 4.40, 6.56, 8.27, 14.34 and 15.24.

[0078] Another embodiment, 50% aqueous alcoholic extract obtained fromthe plant Cuminum cyminum is having HPLC chromatograph peaks with theretention time 4.38, 6.53, 8.25, 10.43, 14.29 and 15.17.

[0079] Still another embodiment, fraction 1 obtained from the plantCuminum cyminum is having HPLC chromatograph peaks with the retentiontime 2.52, 3.59, 6.25, 11.01 and 14.46.

[0080] Still another embodiment, fraction 2 obtained from the plantCuminum cyminum has HPLC chromatograph peaks with the retention time2.43, 4.38 and 6.52.

[0081] Still another embodiment, fraction 3 obtained from the plantCuminum cyminum has HPLC chromatograph peaks with the retention time3.11, 7.75, 10.37, 13.80 and 17.95.

[0082] Still another embodiment, fraction 4 obtained from the plantCuminum cyminum has HPLC chromatograph major peaks with the retentiontime 2.31, 3.14, 5.25, 7.71, 9.63, 13.82 and 17.92.

[0083] Still another embodiment, fraction 5 obtained from the plantCuminum cyminum has HPLC chromatograph major peaks with the retentiontime 2.25, 3.01, 5.21 and 7.69.

EXAMPLES

[0084] The following examples are intended to demonstrate some of thepreferred embodiments and in no way should be construed so as to limitthe scope of the invention. Any person skilled in the art can designmore formulations, which may be considered as part of the presentinvention.

Example 1

[0085] Preparation of colourless, non-pungent piperine by a novelprocess as already claimed in IP 1726890 and further modified asfollows: Commercially available Piper nigrum or Piper longum or theiroleoresins have been used as the source material. 20 kg long pepperoleoresin is extracted with chlorinated solvents like CHCl₃, CH₂Cl₂,C₂H₄Cl₂ (25 litre) for six hours or 20 kg black pepper powder isSoxhletted with toluene for 8 hours. The extracts are concentrated todryness under reduced pressure and dissolved in ethanol at 78° C. Theethanolic solution is adsorbed over neutral Al₂O₃ and packed in a glasscolumn. Elution is carried out with CHCl₃:EtOH (9:1) and the eluate isconcentrated to dryness and dissolved in minimum quantity of ethanol.The solution is treated with activated charcoal and filtered through acelite bed. The filtrate is concentrated to saturation point, cooledwhen colourless crystalline precipitate is obtained. The precipitate isseparated by suction filtration and dried.

Example 2

[0086] The specifications of the preferred materials are as under:

[0087] Piperine

[0088] Colour: Colourless, monoclinic prismatic crystals;

[0089] Melting point: 129°-130° C.

[0090] Assay: Minimum 99.1% (LC/MS)

[0091] BE from C. cyminum

[0092] Preparation and fully fingerprinted (HPLC) products appendedherewith as accompanying drawings.

Example 3

[0093] Doses, models/design of experiments and estimation methodology ina typical experiment is given below:

[0094] Doses of Different Bioenhancers Used and the Design of a TypicalExperiment

[0095] 1. Bioenhancers (BEs) from Cuminum cyminum means either theaqueous, or 50% alcoholic extract or fractions No. 1 to 5.

[0096] 2. Piperine as Bioenhancer (BE) from Piper species means themolecule with characteristics as described in Example 2 of thisinvention.

[0097] 3. In case of Cuminum cyminum, not withstanding the difference indose of extract or its fraction, the enhancement caused in thebioavailability of the drug with which they are combined remains nearlythe same, because the dose of the fraction used is proportionate to itsconcentration in the extract.

[0098] 4. The doses remained either unchanged or were reduced by 50%even when the bioenhancers (BEs) were used in combination with eachother.

Example 4 Doses

[0099] (i) Bioenhancer (BE) from Cuminum cyminum

[0100] Extract: 16 mg/kg body weight (Rats)

[0101] Fraction No. 1: 02-5 mg/kg body weight (rats)

[0102] Fraction No.2: 13-mg/kg body weight (Rats)

[0103] Fraction No. 3: 8 mg/kg body weight (Rats)

[0104] Fraction No. 4: 5 mg/kg body weight (Rats)

[0105] Fraction No. 5: 2.5 to 5 mg/kg body weight (Rats)

[0106] (ii) Piperine: 8 mg/kg body weight (Rats)

[0107] As an example of an experiment in Rat (fasted):

[0108] Drug: Rifampicin, 40 mg/kg

[0109] BE (Cuminum cyminum): Doses as in Example No. 5 above.

[0110] BE (Cuminum cyminum)+Piperine: Doses as in Example No. 5 above.

[0111] Experimental procedure: Drug alone/or in combination with BE wasadministered to rats as per the following design:

[0112] Group 1: Control

[0113] Group 2: Rifampicin alone

[0114] Group 3: BE alone

[0115] Group 4: Rifampicin+BE (Cuminum cyminum)

[0116] Group 5: Rifampicin+BE (C. cyminum+piperine)

[0117] Blood from control/treated animals at predetermined intervals(0-24 hrs) (Total 14 timings). Rifampicin was extracted from the blood(plasma) using dichloromethane. The concentration of rifampicin in thesamples was determined using HPLC (Model: Shimadzu 1080 BP); PDAdetector; Mobile phase: phosphate buffer: acetonitile (40:60); Flow rate1.0 ml/min. Column RP 18.

[0118] Control and BE only groups were employed to study theinterference of plasma component and the bioenhancer used.

Example 5

[0119] The above methodology was adapted for evaluating thebio-enhancing activity of other drugs, micro nutrients, nutracuticals,nutrients and other herbal products and the enhancing effects aretabulated under each heading. List of drugs, nutraceuticals, herbalformulations cited below as some of the example for the purpose ofpresent invention. A. Drugs Categories Drugs I AntibioticsFluoroquinolones Cipro-, Nor-, P-, and 0-floxacins Macrolides Erythro-,Roxythro-, and Azithromycin Cephalosporins Cefalexin, Cefadroxil,cefatrioxone, Cefixime, Cefpirome, Cefdinir Penicillins Amoxycillin,Cloxacillin Aminoglycosides Amikacin, Kanamycin II. AntifungalFluconazole, Amphotericin B, Ketoconazole, Griseofulvin III. Anti-viralAcyclovir, Zidovudine IV. Anti-cancer Methotrexate, 5-Fluorouracil,Doxorubicin Cisplatin V. Cardiovascular drug Amlodipin, Lisinopril &Atenolol VI. CNS drugs Alprazolam & Haloperidol VII Anti-inflammatory/Diclofenac, Piroxicam, Nimesulide & antiarthritic Rofecoxib (NSAID) VIIIAnti-TB/Antileprosy Rifampicin, Ethionamide, Isoniazid, drugsCycloserine Pyrazinamide, Ethambutol Dapsone IX. Anti histamines/Salbutamol, Theophylline, Bromhexine, respiratory disorders LoratidineX. Corticosteroids Prednisolone, dexamethasone, Betamethasone XI.Immuno-suppressants Cyclosporin A, Tacrolimus, Mycophenolate mofetil XIIAntiulcer Ranitidine, cimetidine, omeprazole

[0120] A. Drug Categories: I. Antibiotics: (a) Fluroquinolones %Enhancement in bioavailability BE from Piperine + Active Drug Cuminumcyminum Piperine as BE molecule Ciprofloxacin 52 40 47 P-floxacin 47 5157 O-floxacin 61 40 73 Norfloxacin Negligible Negligible Negligible

[0121] (b) Macrolides % Enhancement in bioavailability BE fromPiperine + Active Drug Cuminum cyminum Piperine as BE moleculeErythromycin 75 105 95 Roxythromycin 67 95 103 Azithromycin 83 91 97

[0122] (c) Cephalosporins % Enhancement in bioavailability BE fromPiperine + BE from Drug Cuminum cyminum Piperine as BE Cuminum cyminumCefalexin 60 70 75 Cefadroxil 90 86 79 Cefatrioxone Nil Nil Nil CefiximeNil Nil Nil

[0123] (d) Penicillins % Enhancement in bioavailability BE fromPiperine + Active Drug Cuminum cyminum Piperine as BE moleculeAmoxycillin 75 111 98 Cloxacillin 94 68 95

[0124] (e) Aminoglycosides % Enhancement in bioavailability BE fromPiperine + Active Drug Cuminum cyminum Piperine as BE molecule AmikacinNil Negligible Nil Kanamycin 95 65 110

[0125] II. Antifungal % Enhancement in bioavailability BE from Piperineas Piperine + Active Drug Cuminum cyminum BE molecule Fluconazole 170110 126 Amphotericin B Nil Negligible negligible Ketoconazole 136 138156

[0126] III. Anti-viral % Enhancement in bioavailability BE fromPiperine + Active Drug Cuminum cyminum Piperine as BE molecule Acyclovir110 77 98 Zidovudine 330 270 415

[0127] IV. CNS drugs % Enhancement in bioavailability BE from Piperine +Active Drug Cuminum cyminum Piperine as BE molecule Alprazolam 60 98 104Haloperidol Nil Nil Nil

[0128] IV. Anti-cancer % Enhancement in bioavailability BE fromPiperine + Active Drug Cuminum cyminum Piperine as BE molecule.Methotrexate 125 70 30 5-Fluorouracil 335 290  435 Doxorubicin 85 42 103Cisplatin 70 Negligible 79

[0129] VI. Cardiovascular drugs % Enhancement in bioavailability BE fromPiperine + Active Drug Cuminum cyminum Piperine as BE moleculeAmlodipine  55  29 103 Lisinopril  83 110  98 Atenolol Nil NegligibleNegligible Propranolol 135 170 210

[0130] VII. Anti-inflammatory/antiarthritic % Enhancement inbioavailability BE from Piperine + Active Drug Cuminum cyminum Piperineas BE molecule. Diclofenac 65 79 108 Piroxicam 70 92 107 Nimesulide 168110 150 Rofecoxib Negligible Negligible Negligible

[0131] VIII. Anti-TB/Antileprosy drugs % Enhancement in bioavailabilityBE from Piperine + Active Drug Cuminum cyminum Piperine as BE molecule.Rifampicin 250 115 366 Isoniazid Nil 22 Negligible Pyrazinamide Nil 17Nil Ethambutol Nil Nil Nil Dapsone 60 40 75 Ethionamide 78 48 65Cycloserine 89 50 90

[0132] IX. Anti-histamines/respiratory disorders % Enhancement inbioavailability BE from Pipenne + Active Drug Cuminum cyminum. Piperineas BE molecule Salbutamol 110 55 85 Theophylline 87 70 75 Bromhexine 5048 90 Loratidine Nil Nil Nil

[0133] X. Corticosteroids % Enhancement in bioavailability BE fromPiperine + Active Drug Cuminum cyminum. Piperine as BE moleculePrednisolone Nil Nil Nil Dexamethasone 85 56 105 Betamethasone 95 65 82

[0134] XI. Immunosuppressants % Enhancement in bioavailability BE fromPiperine + Active Drug Cuminum cyminum Piperine as BE moleculeCyclosporin A 156 223 275 Tacrolimus 75 105 117 Mycophenolate Nil NilNil Mofeit

[0135] XII. Anti-ulcer % Enhancement in bioavailability BE fromPiperine + BE Drug Cuminum cyminum. Piperine as BE Cuminum cyminum.Ranitidine 117 nil  89 Cimetidine 123 Nil 105 Omeprazole Nil Nil Nil

[0136] B. Nutraceuticals % enhancement in bioavailability BE fromPiperine + Dose Cuminum BE from Category (mg/kg) cyminum Piperine C.cyminum I. Vitamins Vitamin A  1 26 14 18 Vitamin E 40 Nil Nil Nil Vit.B1 10 37 16 33 Vit. B6  0.5 Nil Nil Nil Vit B12  0.1 ug Nil Nil Nil Vit.C 50 Nil Nil Nil Folic acid 50 ug Nil Nil Nil II Antioxidants β-Carotene15 45 34 53 Silymarin  5 32 13 41 Selenium  2 Nil Nil Nil III Naturalherbal products Curcumin 50 39 33 29 Boswellic acids 50 Nil Nil NilRutin 40 Nil 26 22 IV Essential nutritional components Methionine 20 2723 30 Lysine 40 35 31 29 Leucine 50 31 25 32 Valine 25 20 26 24Isoleucine 25 40 18 22 Zinc*  0.1 Negligible Nil Nil Calcium* 30 17Negligible Negligible Glucose 50 16 29 11 Potassium* 25 Nil Nil NilCopper* 30 Nil Nil Nil Iron*  0.5 23 Nil 29

[0137] C. Herbal formulations % Enhancement inbioavailability/bioefficacy Piperine + BE BE from Piperine from DrugCuminum cyminum. as BE C. cyminum Echinacea 72 110  90 Tinosporacordifolia 98 107 152 Picrorrhiza kurroa 78  95 115 Aegles marmelos NilNil Nil Andrographis paniculata 72 Nil  68 Emblica ribes 72 Nil  60Asparagus racemosus 35  47  72 Terminalia chebula Nil Nil Nil Withaniasomnifera 55  52  65 Centella asiatica Nil Nil Nil

[0138] Flow Sheet for Preparation of Extracts of Plant Cuminum Cyminum

[0139] Plant material 3.0 Kg Ground to coarse powder Coarse powder * 1.5Kg coarse powder * 1.5 Kg coarse powder * Aqueous extraction * 50% aq.EtOH * Extracted successively extraction four times with deionised*Extracted successively water by heating on water four times with 50%bath at 98±1C for 2 hrs. aqueous EtOH by each time. percolation for 16hrs. * Total solvent used each time.

[0140] Total solvent used 9.5 L (3.5L+3×2.0L) 9.0L (3.0 L+3×2.0L) Liquidextract Liquid extract Clarified * Clarified, EtOH removed on rotovapourat 60° C., reduced pressure * Freeze dried Dry extract Dry extract(Aqueous) (50% aq. alcoholic) 266 gm (17.73%) 265 gm (17.66%)

[0141] Flow Sheet for Fractionation of Aqueous Extract of CuminumCyminum

[0142] Bioactivity Guided Fractionation of Aqueous Extract was Carriedout as Typical Example by Partitioning with n-BuOH and H₂O (SCHEME 1)and Triturating Another Portion of Extract with 95% EtOH, 50% Aq. EtOH(SCHEME-2)

[0143] The original 50% aq. alcoholic extract of the plant material canalso be fractionated by the same scheme as above.

[0144] The original 50% aq. alcoholic extract of the plant material canalso be fractionated by scheme as mentioned above.

1. A bioenhancing/bioavailability-facilitating composition comprising:i. an effective amount of an extract and/or at least one bioactivefraction from Cuminum cyminum; ii. one or more additive selected fromdrugs, nutrients, vitamins, nutraceuticals, herbal drugs/products, micronutrients, antioxidants along with pharmaceutically acceptableadditives/excipient, and iii. optionally, an effective amount ofpiperine or extract/fraction of piper nigrum or piper longum.
 2. Thecomposition as claimed in claim 1, wherein the amount of Cuminun cyminumextract used is in the range of about 1.0 to 250 mg.
 3. The compositionas claimed in claim 2, wherein the amount of Cuminum cyminum extractused is between 2.0 to 100 mg.
 4. The composition as claimed in claims1, wherein the amount of Cuminum cyminum fraction used is in the rangeof about 0.5 to 75.0 mg irrespective of the amount of additive selectedfrom drugs, nutrients, vitamins, nutraceuticals, herbal drugs/products,micro nutrients and antioxidants.
 5. The composition as claimed inclaims 1, wherein the amount of Cuminum cyminum fraction used is in therange of about 1.0 to 30 mg
 6. The composition as claimed in claims 1,wherein the fraction of Cuminum cyminum is selected from fractionsidentified as 1 to
 5. 7. The composition as claimed in claim 1, whereinthe amount of piperine used is in the range of about 2 to 50 mg, moreparticularly between 5 to 20 mg.
 8. The composition as claimed in claim1, wherein piperine is isolated from piper nigrum, piper longum or itsoleoresin.
 9. The composition as claimed in claim 1, wherein the drugsare selected from the group consisting of antibiotics, antifungal,antiviral, anticancer, cardiovascular, CNS drugs,anti-inflammatory/anti-arthritic, anti-TB/anti-leprosy drugs,anti-histamines/drugs for respiratory disorders, corticosteriods,immuno-suppressants, anti-ulcer drugs and herbal drugs.
 10. Thecomposition as claimed in claim 9, wherein the antibiotic used isselected from the group consisting of quinolones, macrolides,cephalosproins, penicillin and aminoglycosides.
 11. The composition asclaimed in claim 10, wherein the quinolone is selected from the groupconsisting of Ciprofloxacin, Pefloxacin, Ofloxacin and Norfloxacin. 12.The composition as claimed in claim 10, wherein the macrolide isselected from the group consisting of Erythromycin, Roxythromycin andAzithromycin
 13. The composition as claimed in claim 10, wherein thecephalosproins is selected from the group consisting of Cefalexin,cefatrioxone, cefixime, Cefpirome, Cefdinir and Cefadroxil.
 14. Thecomposition as claimed in claim 10, wherein the penicillin is selectedfrom the group consisting of Amoxycillin and Cloxacillin.
 15. Thecomposition as claimed in calim 10, wherein the aminoglycocide isselected from the group consisting of Amikacin and Kanamycin.
 16. Thecomposition as claimed in claim 9, wherein anti-fungal drug is selectedfrom the group consisting of Fluconazole, Amphotericin B, Griseofulvinand Ketoconazole.
 17. The composition as claimed in claim 9, whereinantiviral drug is selected from the group consisting of Acyclovir andZidovudine.
 18. The composition as claimed in claim 9, whereinanticancer drug is selected from the group consisting of Methotrexate,5-Fluorouracil, Doxorubicin, Taxol and Cisplatin.
 19. The composition asclaimed in claim 9, wherein cardiovascular drug is selected from thegroup consisting of Amlodipin, Lisinopril, propranolol and Atenolol. 20.The composition as claimed in claim 9, wherein CNS drugs is selectedfrom the group consisting of Alprazolam and Haloperidol
 21. Thecomposition as claimed in claim 9, whereinanti-inflammatory/anti-arthritic drug is selected from the groupconsisting of Diclofenac, Piroxicam, Nimesulide and Rofecoxib.
 22. Thecomposition as claimed in claim 9, wherein anti-TB/anti-leprosy drug isselected from the group consisting of Rifampicin, Ethionamide,Isoniazid, Cycloserine, Pyrazinamide, Ethambutol and Dapsone
 23. Thecomposition as claimed in claim 9, wherein antihistamine/drugs forrespiratory disorders compound is selected from the group consisting ofSalbutamol, Theophylline, Bromhexine and Loratidine
 24. The compositionas claimed in claim 9, wherein corticosteriod is selected from the groupconsisting of Prednisolone, dexamethasone and Betamethasone
 25. Thecomposition as claimed in claim 9, wherein immuno-supressant is selectedfrom the group consisting of Cyclosporin A, Tacrolimus andMycophenolatemofetil.
 26. The composition as claimed in claim 9, whereinthe anti-ulcer compound is selected from the group consisting ofRantidine, Cimetidine and Omerprazole.
 27. The composition as claimed inclaim 1, wherein the herbal product/drug is selected from Echinacea,Tinospora cordifolia, Picrorrhiza kurroa, Aegles marmelos, Andrographispaniculata, Emblica ribes, Asparagus racemosus, Terminalia chebulaWithania somnifera, Centella asiatica and/or their mixture thereof. 28.The composition as claimed in claim 1, wherein the nutrient is selectedfrom group consisting of sugar, carbohydrates, fats and proteins. 29.The composition as claimed in claim 1, wherein vitamin is selected fromthe group consisting of Vitamin A, Vitamin E, Vitamin B1, Vitamin B6,Vitamin B12, Vitamin C and Folic acid.
 30. The composition as claimed inclaim 1, wherein the antioxidant is selected from the group consistingof β-Carotene, Silymarin, Selenium, Lycopene and Ellagiogallotannins 31.The composition as claimed in claim 1, wherein natural herbal product isselected from the group consisting of Curcumin, Boswellic acids andRutin
 32. The composition as claimed in claim 1, wherein essential micronutrients is selected from the group consisting of Methionine, Lysine,Leucine, Valine, Isoleucine, Zinc, Calcium, Glucose, Potassium, Copperand Iron
 33. The composition as claimed in claim 1, wherein, the plantextract of Cuminum cyminum or its bioactive fraction is extracted fromany plant parts of Cuminum cyminum
 34. The composition as claimed inclaim 1, wherein said composition is administered through oral,parenteral, nasal, inhalation including nebulisers, rectal, vaginal,transdermal and any others suitable routes.
 35. The composition asclaimed in claims 1, wherein the bioenhancing effect of theextracts/fractions of Cuminum cyminum alone or in combination withpipeline is selective in enhancing the bioavailability/bioefficacy of adrug, nutraceutical, and herbal drug/formulation.
 36. The composition asclaimed in claim 1, wherein composition containing Cuminum cyminumextract or its bioactive fraction(s) alone providesbioavailability/bio-enhancing activity in the range of 25-335%
 37. Thecomposition as claimed in claims 1, wherein the composition comprisingcomponents of Cuminum cyminum and piperine exhibit nearly significanti.e. about 25% to highly significant i.e. about 435%bioenhancing/bioavailability activity.
 38. The composition as claimed inclaim 1, wherein said composition-containing piperine and Cuminumcyminum provides further bioavailability/bio-enhancing activity in therange of 10 to 85% more than bioenhancing activity of Cuminum cyminumalone.
 39. The composition as claimed in claim 1, wherein the dosage ofCuminum cyminum extract is in the range of 10 to 75-mg/kg/body weight.40. The composition as claimed in claim 1, wherein the dosage of Cuminumcyminum bioactive fraction(s) is in the range of 2 to 30-mg/kg/bodyweight.
 41. The composition as claimed in claim 1, wherein the dosage ofpiperine is in the range of 0 to 12-mg/kg/body weight.
 42. A process forthe preparation of an aqueous extract, aqueous alcoholic extract andbioactive fraction from the plant Cuminum cyminum, said processcomprises steps of: a) extracting crushed plant material with water or50% aqueous alcoholic solvent at a temperature range of 95-100° C. toobtain aq. extract or aq. alcoholic extract respectively, b) extractinga portion of aq. extrtact of step (a) with n-butanol (n-BuOH),separating the n-butanol layer and a aqueous layer, c) evaporating,freeze drying n-BuOH layer of step (b) to obtain fraction 1, d)evaporating, freeze drying the aq. layer of step (b) to obtain fraction2, e) refluxing another portion of the aq.layer of step (a) withalcohol, pooling the alcohol extract and separating the residue leftover, f) evaporating the pooled alcohol extract of step (e) to obtain aresidue as fraction 3, g) extracting residue of step (e) with 50%aq.alcoholic solvent to obtain 50% aq.alcoholic soluble portion andresidue as fraction 5, and h) evaporating the aqueous alocoholic solubleportion of step (g) to obtain a residue as fraction 4.